X-chromosome genes and coronary heart disease In addition to the obvious phenotypic differences between the sexes, there are gender-specific differences in cognitive, immunological and metabolic functions as well as longevity. The goal of our work is to elucidate the genetic and physiological mechanisms underlying these important but poorly understood differences. It was believed that the genetic contribution to sexual dimorphism begins and ends with the action of genes specifying gonads during early embryonic development. This view assumed that only one X-chromosome was active in females and that the Y-chromosome?s unique contribution consists of testis-determining genes, so that all phenotypic differences occurring subsequent to gonadal development were attributed to sex steroid effects. We now realize that many X-chromosome genes escape inactivation and may contribute to gender differences by dosage effects. We aim to identify and define the function of X-chromosome genes involved in the differential development and function of brain, reproductive, metabolic and immune systems in women and men. The study of monsomy X, or Turner syndrome, provides a unique opportunity to elucidate X-chromosome gene dosage effects. This research will enhance our understanding of disease processes such as the increased susceptibility to autoimmune disease in women and increased risk for coronary disease in men. The idea that estrogen protects women against CHD has recently been refuted by major clinical studies and there is ample evidence exculpating androgens for the increased CHD risk in men. To investigate the potential contribution of X-chromosome gene(s) to the protection from CHD, we examined CHD risk factors in women with Turner Syndrome (TS). TS is characterized by short stature, ovarian dysgenesis, cardiovascular anomalies and premature CHD. To control for the ovarian failure in TS, we compared glucose and lipid metabolism in lean, young women with TS and age- and body composition-matched women with karyotypically normal premature ovarian failure (POF). Body mass and percent adipose tissue are similar in the 2 groups, but total cholesterol, LDL cholesterol and triglycerides are all significantly increased in TS. Moreover, NMR spectroscopy revealed a concentration of smaller, denser HDL and LDL lipid particles in women with TS in contrast to those with karyotypically normal POF. These data show a distinctly atherogenic lipid profile in otherwise healthy, non-obese young women with TS. Diabetes mellitus is another major risk factor for CHD. Reports on diabetes mellitus (DM) in TS have varied widely, with some studies indicating an increased prevalence up to 40% and others finding no increase over normal populations. We have now shown that while most girls and women with TS have normal fasting glucose and insulin, the glycemic response to a glucose challenge is dramatically abnormal and consistent with diabetes in about 40%, and is significantly above the POF control group in all women with TS. Interestingly, the glucose intolerance in these young lean women and girls with TS is not explained by insulin resistance, but by a novel insulin secretory defect. The insulin response to an oral or IV glucose challenge is significantly lower than POF or normal controls in all women with TS. It thus appears that the Turner metabolic syndrome is is a distinct entity characterized by decreased insulin secretion, reminiscent of transcription factor mature onset diabetes of the young (MODY) syndromes, caused by haploinsufficiency for autosomal genes involved in pancreatic development, suggesting that haploinsufficiency for unknown X-chromosome gene(s) impairs beta cell function and predisposes to diabetes mellitus in TS. Our new findings implicating haploinsufficiency for X-chromosome genes in dyslipidemia and diabetes explain the increased risk for CHD in women with TS, and may also contribute to the increased risk for CHD among normal XY men compared to women. The identification of these genes clearly is of great clinical importance. We are using traditional approaches through genotype-phenotype evaluations in subjects with informative X-chromosome deletions or rearrangements, and novel bioinformatic strategies to identify functionally interesting X-chromosome sequences likely to escape inactivation. The role of androgen in protection of mammary gland The normal ovary produces abundant quantities of testosterone (T) in addition to estradiol, but usual hormone replacement treatment (HRT) for ovarian failure consists of estrogen(E) and progesterone(P). The risk of breast cancer is increased in post-menopausal women with such treatment, however. We have proposed that an important role for endogenous androgen in women is to protect the mammary gland from unopposed estrogenic stimulation. We showed that an androgen antagonist doubled mammary epithelial proliferation (MEP)in normal monkeys suggesting that endogenous androgens normally inhibit MEP. In an ovx monkey model, we showed that mammary epithelial proliferation (MEP) was increased by ~4-fold by E and E/P treatment, but was no different from vehicle-treated control with E/T treatment. We also showed that estrogen receptor (ER) alpha is down-regulated and ER beta up-regulated by testosterone. Since the alpha isoform promotes proliferation and MYC expression, while the beta isoform does not, the dramatic alteration in ER alpha/beta ratio may be key to T' inhibition of E-induced proliferation. These observations suggest that endogenous androgens normally limit mammary epithelial proliferation and that androgen supplementation of estrogen therapy may reduce estrogen-induced proliferation and breast cancer risk. These considerations are of crucial importance for young women with ovarian failure who face a long period of HRT, as well as those post-menopausal women who need HRT. IGF1 and the brain We have previously shown that endogenous brain IGF1 serves an insulin-like role in promoting neuronal glucose utilization and hence growth during postnatal development. We have shown that brain growth in Igf1 null mice falls behind that of normal littermates by almost 40% during the postnatal period. Over this past year, we have shown that neuronal numbers are preserved in the Igf1 null neocortex, but that pyramidal neuron soma size is reduced by ~10% and Golgi staining showed a significant reduction in pyramidal dendritic length and complexity. In addition, the density of dendritic spines and presumably synaptic contacts was reduced by 16% (P = 0.002) in the Igf1 null brain. Taken together, these findings illustrate multifaceted roles for IGF1 in post-natal brain development, and explain why individuals with IGF1 gene deletions demonstrate mental retardation in addition to short stature. We have recently shown that a ketogenic diet (KD) regulates brain IGF and IGF receptor expression in a rat model for epilepsy. The KD is a high-fat, low-carbohydrate diet that is used for treating refractory epilepsy in children. Despite its long history of clinical use, it is still not clear how KD affects the brain and what mechanism(s) underlie its seizure-suppressive action. Reduction in brain energy supply, e.g., from systemic hypoglycemia or from reduced brain GLUT1 expression induces seizure activity by impairing the ability of neurons to stabilize membrane potential. Since IGF1 is a key regulator of brain glucose metabolism,we propose that KD-indued enhanced IGF1 activity improves energy utilization and thus promotes protection from seizures.